báo cáo khoa học: " Genetic control of mammalian T-cell proliferation with a synthetic RNA regulatory system - illusion or reality?" pps

In the past two decades, molecular biology research has
revealed the intimate mechanisms of epidemiologically
significant diseases, such as cancer, infections and
immunological disorders. As the next step beyond
seeking the mechanisms involved, scientists are now
increasingly making it possible to regulate human bio-
logical reactions. In recent years, there have been break-
throughs in genetic engineering related to the inventory
and methods necessary to physically construct and
assemble biomolecular parts, such as synthetic RNA-
based regulatory systems [1]. Synthetic biology relies on
the engineering of biological systems that perform
human-defined functions and on the synthesis of
complex, biologically based systems that show functions
that do not exist in nature. Despite the possible
advantages for clinical applications, more work remains
to be done to elucidate the principles of biological
design, and to overcome the scientific and technical
challenges in designing and building more effective
systems that are harmless to humans and therefore
useful for clinical applications.
A recent study by Chen et al. [2] has produced a signi fi-
cant advance in solving such issues and therefore
potentially bridging the gap between the bench and the
bedside for synthetic RNA-based regulatory systems. e
authors [2] developed a modular device composed of a
sensor (an aptamer) and a gene-regulatory component (a
hammerhead ribozyme) and tested its ability to affect the
expression of cytokines important for the function of T-
lymphocytes in mouse and human systems.
Why is this work [2] significant? First, it represents the

logical continuation of years of experimental work
performed by the same group, coming from a team that
understands the way a synthetic RNA-based regulatory
system works and its immediate practical applications. In
fact, in a previous study [3], also published in Proceedings
of the National Academy of Sciences of the United States
of America, the authors were the first to develop and set
up universal RNA-based regulatory platforms, called
ribozyme switches, by using engineering design princi-
ples. In the present report [2], the authors expanded the
advantages of such biomodular platforms to a broader
range of applications. ey were able to do so by the
reliable de novo construction of modular, portable and
scalable control systems that can achieve flexible regu la-
tory properties, such as up- and down-regulation of
target expression levels and tuning of regulatory res-
ponses to fit application-specific performance requirements.
Second, the authors [2] applied the synthetic RNA
regulatory device to a significant medical issue, the use of
adoptive cell transfer (ACT) [4]. e ACT strategy uses
T-cell-based cytotoxic responses to attack malignant cells
(or any other types of abnormal cells) that escape the
body’s natural surveillance by using T cells that have a
natural or genetically engineered reactivity to a patient’s
cancer cells. For this purpose, T cells have first to be
Abstract
Synthetic RNA-based regulatory systems are used
to program higher-level biological functions that
could be exploited, among many applications, for in
vivo diagnostic and therapeutic applications. Chen

and colleagues have recently reported a signicant
technological advance by producing an RNA modular
device based on a hammerhead ribozyme and
successfully tested its ability to control the proliferation
of mammalian T lymphocytes. Like all exciting research,
this work raises a lot of signicant questions. How
quickly will such knowledge be translated into clinical
practice? How ecient will this system be in human
clinical trials involving adaptive T-cell therapy? We
discuss the possible advantages of using such new
technologies for specic therapeutic applications.
© 2010 BioMed Central Ltd
Genetic control of mammalian T-cell proliferation
with a synthetic RNA regulatory system - illusion or
reality?
Sang Kil Lee
1,2
and George A Calin
1
*
CO M M E N TA R Y
*Correspondence:
1
RNA interference and non-coding RNA Center and the Department of
Experimental Therapeutics, University of Texas, MD Anderson Cancer Center,
Houston, TX 77030, USA
Full list of author information is available at the end of the article
Lee and Calin Genome Medicine 2010, 2:77
/>© 2010 BioMed Central Ltd
naturally or genetically engineered to react against a

tumor-specific antigen, then expanded and made more
effective in vitro, and finally adoptively transferred into a
cancer patient. However, the clinical efficacy of ACT, so
far, has been limited. ere are many reasons for this, and
insufficient persistence and reactivation of infused T cells
are among the main ones. Conventional strategies for
enhancing the persistence of transferred T cells include
ablation of all white blood cells (myeloablative methods),
such as total body irradiation and administration of toxic
levels of interleukin (IL)-2. However, myeloablation is
associated with considerable morbidity, caused by
decreased immune response and increased risk of infec-
tion [5]. erefore, safer and more effective thera peutic
strategies are yet to be discovered.
Chen et al. [2] report on a synthetic RNA regulatory
system, which marks a new era in adoptive T-cell therapy
because of the increase in the amount and survival of
infused T cells found with this system. eir system for
the control of mammalian T-cell proliferation is based on
a platform of assembled RNA devices formed by a
modular sensor (aptamer) and a gene-regulatory
(hammer head ribozyme) component. is device con-
verts a small-molecule input to an increased gene
expression output, in this particular case cytokine
production. In more detail, the authors [2] fused a
theophylline ribozyme switch to the 3’ untranslated
region of a tri-functional transgene (cd19-tk-t2a-il15)
encoding IL-15 (potent survival/proliferative cytokine of
T cells), mutant HSV-1 thymidine kinase (acting as a
reporter and as a suicide protein in the presence of

ganciclovir) and CD19 (a marker for fluorescence-activated
cell sorting and immuno magnetic selection). Using this
system, they could strictly measure (by monitoring the
expression of CD19) and control (by modulating the
levels of the input molecule) the biological response
(cell proliferation/viability).
In addition to all the in vitro evidence, the authors [2]
demonstrated that this system worked in vivo and
effectively modulated the T-cell growth rate in mice in
response to theophylline administration. e growth rate
was increased to 32% in the presence of theophylline over
a 14 day study in mice. ey further investigated its
possible clinical application by transducing primary
human central memory T cells with this system. In vitro
results showed that the population of live central memory
T cells increased by 24% and that apoptotic cell popu-
lation was decreased by 54% in the theophylline-
responsive system [2].
Finally, the presented gene regulatory system [2]
showed significant advantages over available gene regu-
la tory techniques (synthetic inducible promoters); in
par ticular, it provides a wide range of flexibility for
clinical settings. Firstly, the ribozyme switches can be
easily programmed to respond to different drug
molecules. Secondly, the system can be stringently
controlled and finely tuned by adding additional drug-
responsive ribozyme switches (up to four), therefore
achieving lower basal gene expression levels. irdly,
this system shows tight drug-mediated regulation of
growth over an extended time period. Taking all these

features into consideration, it is feasible that this new
synthetic RNA-based regulatory system could have
straightforward clinical utility.
It is likely that combining this new modular device
framework with upcoming advances in synthetic biology
will strongly support the tailoring of RNA-based
regulatory systems to diverse applications in various
clinical and laboratory environments. Yet applying these
RNA-based regulatory systems in clinical practice may
still require more time. In addition, there are many other
factors that limit the use of adoptive T-cell therapy for
cancer. For example, the failure of adoptive immuno-
therapy against cancers lies in the absence of tumor-
specific sources of T cells [6]. If such obstacles are not
overcome, efficacy of these systems will be significantly
limited in clinical practice. Also, recent data support the
combined roles of protein-coding genes and non-coding
RNAs, such as microRNAs, in the pathogenesis of
frequent diseases (such as cancer, immune and cardiac
disorders) [7]. One question for the future is whether
such devices can be adapted for the regulation of the
functions of non-coding RNAs and microRNAs. e
published research is good news, but it would be better
to hold our cheers until the clinical trials are successfully
completed, which we hope will be in the near future.
Abbreviations
ACT, adoptive cell transfer.
Competing interests
The authors declare that they have no competing interests.
Authors’ contributions

Both authors wrote the article.
Author information
GAC received his MD and PhD at Carol Davila University of Medicine in
Bucharest, Romania. After working on cytogenetics as an undergraduate
student with Dragos Stefanescu in Bucharest, he completed training in
cancer genomics in Massimo Negrini’s laboratory at the University of
Ferrara, Italy. In 2000 he became a postdoctoral fellow at the Kimmel
Cancer Center in Philadelphia, in Carlo Croce’s laboratory. Since July 2007
he has been an associate professor in Experimental Therapeutics at the
MD Anderson Cancer Center and studies the roles of microRNAs and
other non-coding RNAs in cancer initiation and progression, as well as the
mechanisms of cancer predisposition, and explores new RNA therapeutic
options for cancer patients. SKL graduated from Yonsei University Medical
School, Seoul, South Korea with an MD and PhD and is an assistant
professor in the Department of Gastroenterology, Severance Hospital,
Seoul. His primary clinical focus is treating colon and gastric cancers. The
focus of his scientic research is understanding the roles of non-coding
RNAs in gastrointestinal cancers. In March 2009, he began working in GAC’s
laboratory studying the roles of non-coding RNAs, including microRNAs in
the initiation and development of gastrointestinal cancers, as well as the
identication of new non-coding RNA biomarkers.
Lee and Calin Genome Medicine 2010, 2:77
/>Page 2 of 3
Acknowledgements
We thank Milena Nicoloso for critically reading this manuscript. GAC is
supported as a Fellow at The University of Texas MD Anderson Research Trust,
as a Fellow of The University of Texas System Regents Research Scholar and by
the CLL Global Research Foundation. Work in GAC’s laboratory is supported
in part by NIH, by DOD, by Developmental Research Awards in Breast Cancer,
Ovarian Cancer and Leukemia SPOREs, and by a 2009 Seena Magowitz

Pancreatic Cancer Action Network AACR Pilot Grant.
Author details
1
RNA interference and non-coding RNA Center and the Department of
Experimental Therapeutics, University of Texas, MD Anderson Cancer Center,
Houston, TX 77030, USA.
2
Institute of Gastroenterology, Department of
Internal Medicine, Yonsei University College of Medicine, 250 Seongsanno,
Seodaemun-gu, Seoul 120-752, South Korea.
Published: 15 October 2010
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doi:10.1186/gm198
Cite this article as: Lee SK, Calin GA: Genetic control of mammalian T-cell
proliferation with a synthetic RNA regulatory system - illusion or reality?
Genome Medicine 2010, 2:77.
Lee and Calin Genome Medicine 2010, 2:77
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